Vol.

11, 105–111, January 2002 Cancer Epidemiology, Biomarkers & Prevention 105

Metabolism of the Cancer Chemopreventive Agent Curcumin in

Human and Rat Intestine

Christopher R. Ireson, Donald J. L. Jones, Equine alcohol dehydrogenase catalyzed the reduction of
Samantha Orr, Michael W. H. Coughtrie, curcumin to hexahydrocurcumin. The results show that
David J. Boocock, Marion L. Williams, Peter B. Farmer, curcumin undergoes extensive metabolic conjugation and
William P. Steward, and Andreas J. Gescher1 reduction in the gastrointestinal tract and that there is
Medical Research Council Toxicology Unit, University of Leicester, Leicester more metabolism in human than in rat intestinal tissue.
LE1 9HN, United Kingdom [C. R. I., D. J. L. J., D. J. B., P. B. F., A. J. G.]; The pharmacological implications of the intestinal
Human Tissue Bank and School of Pharmacy and Pharmaceutical Sciences, De metabolism of curcumin should be taken into account in
Montfort University, Leicester LE1 9BH, United Kingdom [S. O.]; Department
of Molecular and Cellular Pathology, University of Dundee, Ninewells
the design of future chemoprevention trials of this dietary
Hospital & Medical School, Dundee DD1 9SY, United Kingdom constituent.
[M. W. H. C.]; and Department of Oncology, University of Leicester, Leicester
LE1 5WW, United Kingdom [M. L. W., W. P. S.]
Introduction
Curcumin [1,7-bis(4-hydroxy-3-methoxyphenyl)-1,6-heptadiene-
Abstract 3,5-dione] is the major yellow pigment extracted from turmeric, a
Curcumin, the yellow pigment in turmeric, prevents commonly used spice, derived from the rhizome of the herb
malignancies in the intestinal tract of rodents. It is Curcuma longa Linn. In the Indian subcontinent and Southeast
under clinical evaluation as a potential colon cancer Asia, turmeric has traditionally been used as a treatment for in-
chemopreventive agent. The systemic bioavailability of flammation, skin wounds, and tumors. Clinical activity of curcu-
curcumin is low, perhaps attributable, at least in part, to min has yet to be confirmed; however, in preclinical animal
metabolism. Indirect evidence suggests that curcumin is models, curcumin has shown cancer chemopreventive, antineo-
metabolized in the intestinal tract. To investigate this plastic, and anti-inflammatory properties (for review, see Ref. 1).
notion further, we explored curcumin metabolism in Especially interesting is its ability to prevent the formation of
subcellular fractions of human and rat intestinal tissue, carcinogen-induced intestinal premalignant lesions and malignan-
compared it with metabolism in the corresponding cies in rats (2, 3) and in the multiple intestinal neoplasia (Min/⫹)
hepatic fractions, and studied curcumin metabolism mouse (4), a genetic model of the human disease familial ade-
in situ in intact rat intestinal sacs. Analysis by high- nomatous polyposis. Curcumin acts as a scavenger of oxygen
performance liquid chromatography, with detection at species, such as hydroxyl radical, superoxide anion, and singlet
420 or 280 nm, permitted characterization of curcumin oxygen (5–9), and it interferes with lipid peroxidation (10 –12).
conjugates and reduction products. Chromatographic Curcumin suppresses a number of key elements in cellular signal
inferences were corroborated by mass spectrometry. transduction pathways pertinent to growth, differentiation, and
Curcumin glucuronide was identified in intestinal malignant transformation. Among signaling events inhibited by
and hepatic microsomes, and curcumin sulfate, curcumin are protein kinases (13), c-Jun/AP-1 activation (14),
tetrahydrocurcumin, and hexahydrocurcumin were found prostaglandin biosynthesis (15), and activity and expression of the
as curcumin metabolites in intestinal and hepatic cytosol enzyme cyclooxygenase-2 (16). This latter property is probably
from humans and rats. The extent of curcumin mediated via the ability of curcumin to block activation of the
conjugation was much greater in intestinal fractions from transcription factor NF-␬B at the level of the NF-␬B-inducing
humans than in those from rats, whereas curcumin kinase/IKK␣/␤ signaling complex (17). In rodents, curcumin dem-
conjugation was less extensive in hepatic fractions from onstrates poor systemic bioavailability after p.o. dosing (18),
humans than in those from rats. The curcumin-reducing which may be related to its inadequate absorption and avid me-
ability of cytosol from human intestinal and liver tissue tabolism. Curcumin bioavailability may also be poor in humans, as
exceeded that observed with the corresponding rat tissue borne out by a recent pilot study of a standardized Curcuma extract
by factors of 18 and 5, respectively. Curcumin sulfate in colorectal cancer patients (19). After p.o. dosing, curcumin
was identified in incubations of curcumin with intact undergoes metabolic O-conjugation to curcumin glucuronide and
rat gut sacs. Curcumin was sulfated by human phenol curcumin sulfate and bioreduction to tetrahydrocurcumin, hexa-
sulfotransferase isoenzymes SULT1A1 and SULT1A3. hydrocurcumin, and hexahydrocurcuminol (Fig. 1) in rats and
mice in vivo (18, 20, 21) and in suspensions of human and rat
hepatocytes (18). Products of curcumin reduction are also subject
to glucuronidation (20). Certain curcumin metabolites, such as tet-
Received 7/6/01; revised 10/26/01; accepted 10/31/01. rahydrocurcumin, possess anti-inflammatory (11) and antioxidant
The costs of publication of this article were defrayed in part by the payment of activities (22, 23) similar to those of their metabolic progenitor. It
page charges. This article must therefore be hereby marked advertisement in has been suggested that the intestinal tract plays an important role
accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
1
To whom requests for reprints should be addressed, at the MRCouncil Toxi-
in the metabolic disposition of curcumin. The evidence supporting
cology Unit, University of Leicester, Leicester LE1 9HN, United Kingdom. this notion is based predominantly on experiments in which
Phone: 44 (0) 116 2525618; Fax: 44 (0) 116 2525616; E-mail: ag15:@le.ac.uk. [3H]labeled curcumin was incubated with inverted rat gut sacs, and

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106 Intestinal Metabolism of Curcumin

which inhibits SULT enzymes (25). To remove this contami-

nant from PAPS, the commercial product was purified by
HPLC, essentially as described previously (26) using a Varian
Prostar (310 model) solvent delivery system coupled to an
octadecyl silan reversed phase C18 column (4.6 ⫻ 250 mm;
Beckman) and a UV-visible detector. Aliquots (100 ␮l) of the
PAPS solution (4 mM) were injected into the column. The
eluant flow rate was 1.3 ml/min. Eluant was collected on dry
ice, and PAPS was concentrated by rotary evaporation (4 min,
room temperature). The collected PAPS was ⬎99% pure by
HPLC analysis.
Preparation of Rat and Human Intestinal and Hepatic Mi-
crosomes and Cytosol. Experiments using animals were con-
ducted as stipulated by Project License 80/1250 granted to the
Fig. 1. Chemical structures of curcumin and products of its bioreduction. MRC Toxicology Unit by the United Kingdom Home Office,
and the experimental design was vetted and approved by the
Leicester University Ethical Committee for Animal Experimen-
tation. Male F344 rats were subjected to terminal anesthesia
biotransformation was deduced from the disappearance of radio- (halothane/nitrous oxide), and blood was removed by cardiac
activity associated with the parent compound (24). Metabolites of puncture. Liver and intestine were removed and snap frozen in
curcumin have hitherto not been unambiguously identified in gut liquid nitrogen. Human tissue was obtained from the United
tissue. Therefore, we tested the hypothesis that curcumin is met- Kingdom Human Tissue Bank (Leicester, United Kingdom);
abolically conjugated and/or reduced in intestinal tissue. This healthy tissues had been resected from 6 Caucasian patients
hypothesis was tested using two experimental settings: cytosolic (livers from 1 male, who was 4 years of age, and 2 females, 30
and microsomal fractions of intestinal tissue obtained from hu- and 51 years of age; intestine from 3 females, who were 29, 54,
mans and rats and suspensions of intact rat gut sacs under the
and 56 years of age). Patients had not received medication
conditions used to make the original observations regarding the
known to interfere with xenobiotic metabolism activity. Human
intestinal metabolism of [3H]curcumin (24). To be able to obtain
and rat intestinal or hepatic tissue was defrosted and weighed,
an indication of the quantitative importance of the contribution of
and microsomes and cytosol were prepared as described pre-
intestinal tissue to the overall metabolism of curcumin in the
viously (27). Human intestinal tissue originated from the jeju-
organism, the extent of its biotransformation in intestinal tissue
nal area of the intestine, and rat intestinal tissue came from the
fractions was compared with that in analogous liver fractions.
jejunum and colon.
Because the knowledge of enzymatic details of curcumin biotrans-
formation is only rudimentary, we also addressed the question as Metabolism of Curcumin by Intestinal and Hepatic Subcel-
to which enzymes may be involved in the metabolic generation of lular Fractions. To study curcumin conjugation, substrate
curcumin sulfate and hexahydrocurcumin, two major curcumin (100 ␮M) was incubated for 1 h with hepatic or intestinal
metabolites that we identified in intestinal tissue. Overall, the cytosol or microsomes (1 mg of cytosolic or microsomal pro-
experiments were designed to contribute to the body of knowledge tein/ml) in phosphate buffer (0.01 M) at 37°C. Cytosolic or
that will ultimately help rationalize the design of future chemo- microsomal protein was quantitated using the Bio-Rad protein
prevention trials of curcumin. assay kit (Bio-Rad Laboratories GmbH, Munich, Germany).
Incubations to study glucuronidation included microsomes, uri-
dine 5⬘-diphosphoglucuronic acid (3 mM), magnesium chloride
Materials and Methods (5 mM), and Triton X-100 (0.01%) in phosphate buffer at pH
Chemicals and Reagents. The following chemicals and re- 7.4. Incubations to study sulfation included cytosol with PAPS
agents were purchased from the suppliers listed: curcumin, uridine (0.4 mM) and mercaptoethanol (5 mM) in phosphate buffer at
5⬘-diphosphoglucuronic acid, magnesium chloride, PAPS,2 aden- pH 8.4. Curcumin sulfate and curcumin glucuronide were quan-
osine 3⬘-5⬘diphosphate, NADPH, equine alcohol dehydrogenase, titated with the help of a calibration curve established using
and Triton X-100: Sigma Chemical Co.-Aldrich Comp., Ltd. curcumin. In orientation experiments, curcumin glucuronide
(Poole, Dorset, United Kingdom). Authentic curcumin glucuro- and sulfate generation were found to be linear for ⱕ30 min,
nide, curcumin sulfate, tetrahydrocurcumin, and hexahydrocur- after which the rate of conjugate formation declined. Therefore,
cumin were synthesized as described (18), and the latter two were the values shown in Table 1 are amounts generated per h. To
provided by Dr. W. Wang (Phytopharm plc, Godmanchester, study curcumin bioreduction, substrate (100 ␮M) was incubated
United Kingdom). In experiments in which the metabolism of with cytosol or microsomes and NADPH (1 mM) in phosphate
curcumin or hexahydrocurcumin was studied in incubations with buffer (10 mM, pH 7.4) in a final volume of 0.5 ml at 37°C.
tissue fractions, gut sacs, or enzymes, substrates were dissolved in Incubation time was 90 min, during which generation of hexa-
DMSO, and an aliquot of 5 ␮l was added to incubate to furnish a hydrocurcumin was linear. The amount of hexahydrocurcumin
final substrate concentration of 100 ␮M. generated was assessed using a calibration curve with authentic
Purification of PAPS. The purity of the purchased PAPS was hexahydrocurcumin. In control experiments, substrate and re-
determined by HPLC to be only 80%. Commercial PAPS is action components were incubated with microsomal or cytoso-
often contaminated with phosphoadenosine 5⬘-phosphate, lic fractions in which enzymes had been inactivated by expo-
sure to boiling water for 10 min. Reactions were terminated by
cooling incubate samples to ⫺80°C.
2
The abbreviations used are: PAPS, 3⬘-phosphoadenosine 5⬘-phosphosulfate;
Metabolism of Curcumin by Intact Rat Intestine. The jeju-
MRC, Medical Research Council; HPLC, high-performance liquid chromatogra- nal section of the small intestine of terminally anesthetized
phy; SULT, sulfotransferase. male F344 rats (180 grams) was excised. Gut content was

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 Cancer Epidemiology, Biomarkers & Prevention 107

Table 1 Metabolites of curcumin in cytosolic and microsomal fractions of liver and intestine of humans and rats

Intestine (nmol/mg protein) Liver (nmol/mg protein)

Metabolite Cytosol Microsomes Cytosol Microsomes
a b
Rat Human Rat Human Rat Human Rat Human

Hexahydrocurcumin 8⫾3 142 ⫾ 92 N.D.c N.D. 25 ⫾ 12 121 ⫾ 116 N.D. N.D.

Curcumin sulfate 25 ⫾ 3 103 ⫾ 23 N.D. N.D. 182 ⫾ 39 39 ⫾ 8 N.D. N.D.
Curcumin glucuronide N.D. N.D. 16 ⫾ 1 255 ⫾ 229 N.D. N.D. 980 ⫾ 150 94 ⫾ 30
a
Mean ⫾ SD (n ⫽ 4).
b
Mean ⫾ SD (n ⫽ 3).
c
N.D., not detected.

removed by flushing with 0.9% (w/v) sodium chloride solution. Results

Everted gut sacs of ⬃8-cm length were prepared as described Curcumin Metabolism in Subcellular Fractions of Intestine
previously (28) using a glass rod. Sacs were suspended in an and Liver. Curcumin was incubated with intestinal and he-
Ussing chamber in Krebs-Ringer phosphate buffer (10 ml), patic cytosol and microsomes obtained from humans and rats.
containing glucose (10 mM) and curcumin to give a final con- Extracts of the incubation mixtures were analyzed by HPLC for
centration of 100 ␮M. Buffer with substrate was added to the curcumin, and its metabolites were analyzed with detection by
lumen of the gut to ensure full extension. Gut sacs were incu- UV/visible spectrophotometry at 420 and 280 nm and by mass
bated with curcumin for 1 h under a continual stream of spectrometry in the selected ion monitoring mode. Spectropho-
carbogen at 37°C. tometric analysis at 420 nm indicated the presence of mole-
Metabolism of Curcumin by Isolated Enzymes and Western cules, such as curcumin sulfate and curcumin glucuronide,
Analysis. Curcumin (100 ␮M) was incubated with recombinant containing the intact yellow-colored diferuloylmethane struc-
SULT1A1 and 1A3 (10 ␮g/ml) obtained as described previ- ture, whereas detection at 280 nm also allowed characterization
ously (29) or with equine alcohol dehydrogenase (10 units/ml) of molecules, such as tetrahydrocurcumin and hexahydrocur-
in phosphate buffer (0.01 M, pH 8.4 for sulfation, pH 7.4 for cumin, generated from curcumin by reductive destruction of the
reduction); the final volume was 0.5 ml. Incubations contained chromophoric diarylheptatrienone chain. Metabolite analysis
PAPS (0.4 mM, sulfation) or NADPH (10 mM, reduction) and was aided by chromatographic comparison with authentic ref-
were conducted at 37°C for 1 h. Metabolites were extracted as erence compounds. Analysis of incubates of curcumin with
described above. SULT1A1/1A3 primary antibodies were pre- human and rat intestinal cytosol (Fig. 2) yielded curcumin
pared and used for Western blotting as described previously sulfate and hexahydrocurcumin. Both species were character-
(29). ized mass spectrometrically by molecular ions of m/z ⫽ 447
HPLC Analysis. After acidification with acetate buffer (1 M, and 373, respectively. Fig. 3A shows the selected ion chromat-
pH 4.6), samples were extracted twice with ethyl acetate:pro- ogram of curcumin sulfate generated in human gut cytosol.
pan-2-ol (9:1), and mixtures were centrifuged (2800 ⫻ g, 4°C, There was also evidence of the presence of tetrahydrocurcumin
15 min). 5,10,15,20-Tetra-(m-hydroxylphenyl)-chlorine served as adjudged by mass spectrometry (molecular ion m/z ⫽ 371).
as internal standard. The organic layers were removed and On UV-spectrophotometric detection, authentic tetrahydrocur-
combined, and solvent was evaporated to dryness under nitro- cumin gave a broad shoulder with a nonsymmetrical peak,
gen. A reversed-phase HPLC method described before (18) was probably the corollary of an unstable equilibrium of stereoiso-
used to determine the quantity of curcumin and its metabolites. mers, which are possible for the 1,7-diarylhepta-(3,4-ene)-5-
HPLC analysis was performed on a Varian Prostar (230 model) one structure (Fig. 1). Furthermore, a small peak in the cyto-
solvent delivery system coupled to a UV-visible detector (310 solic extracts eluted at the retention time of authentic
model) and autosampler (model 410) and a Symmetry Shield hexahydrocurcuminol (retention time: 22 min), but this species
RP 18 column (150 ⫻ 3.9 mm, Waters). Detection of curcumin, eluded conclusive identification. HPLC analysis of extracts of
curcumin sulfate, and curcumin glucuronide was achieved at incubates of curcumin with human or rat intestinal microsomes
420 nm, whereas tetrahydrocurcumin, hexahydrocurcumin, and (Fig. 2) afforded a peak consistent with curcumin glucuronide,
hexahydrocurcuminol were analyzed at 280 nm. The extraction as adjudged by its chromatographic properties and its molecular
efficiency for hexahydrocurcumin from incubations with tissue ion of m/z ⫽ 543. Microsomes did not generate detectable
fractions was 92 ⫾ 7% (mean ⫾SD, n ⫽ 6), which for curcu- levels of products of curcumin reduction. Results qualitatively
min was very similar (18). In contrast, the extraction efficien- similar to those shown in Fig. 2 were obtained with hepatic
cies for curcumin glucuronide and curcumin sulfate were only cytosol and microsomes (results not shown).
50 ⫾ 9%. Quantitation of Curcumin Metabolites. Quantitative analy-
Mass Spectrometry. Mass spectrometry was performed as sis revealed considerable differences in curcumin metabolite
described before (18) using a Quattro Bio-Q tandem quadruple generation between human and rat tissue and between gut and
mass spectrometer upgraded to Quattro MK II specifications liver when values were normalized to cytosolic or microsomal
(Micromass, Altrincham, Cheshire, United Kingdom) with a protein content (Table 1). The extent of sulfation of curcumin
pneumatically assisted electrospray interface. Samples were in the cytosol of human intestinal tissue was four times that in
analyzed in negative ion mode. The temperature was main- rat intestine, whereas in human liver cytosol, it was only a fifth
tained at 120°C, the operating voltage of the electrospray cap- of that observed in rat liver cytosol. Curcumin sulfation was
illary was 3.88 kV, and the cone voltage was 32 V. HPLC 3-fold higher in cytosol from human intestine than in that from
conditions used for the on-line HPLC-mass spectrometric anal- human liver, whereas in the rat, intestinal sulfation was only a
yses were as described before (18). seventh of that in the liver. Microsomal metabolism of curcu-

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108 Intestinal Metabolism of Curcumin

Fig. 2. High-performance liquid chromatograms of extracts of incubations of curcumin (100 ␮M) with cytosol (A and C) and microsomes (B) from human intestinal tissue
and with cytosol (D and F) and microsomes (E) from rat intestinal tissue. Incubation periods were 90 min for metabolic reduction (A and D) and 60 min for conjugation
(B, C, E, and F). Chromatographic analysis was conducted with detection at 280 (A and D) and 420 nm (B, C, E, and F). The identity of the peaks was established by
cochromatography and mass spectrometry as curcumin (peak 3), hexahydrocurcumin (1), tetrahydrocurcumin (2), curcumin glucuronide (4), and curcumin sulfate (5). The
prominent peak labeled “is ” (retention time: 51 min) was caused by the internal standard 5,10,15,20-tetra-(m-hydroxylphenyl)-chlorine. Note that commercially available
curcumin contains 15% desmethoxycurcumin and 5% bisdesmethoxycurcumin, which furnished two small peaks just beyond curcumin (see especially in A and D). Removal
of the methoxy moieties from the curcumin molecule appears to render its phenolic groups more resistant against metabolic conjugation, and bisdesmethoxycurcumin was
not a substrate of the conjugating enzymes. AU, absorbance units. For details of incubation, extraction, and HPLC analysis, see “Materials and Methods.” The
chromatograms are representative of analyses conducted with fractions or tissues from three humans or four rats.

higher in human intestinal cytosol and five times more abun-

dant in human liver cytosol compared with the corresponding
values in rats. Production of hexahydrocurcumin occurred to a
similar extent in the cytosol of human intestinal and hepatic
tissue. In the rat, reduction of hexahydrocurcumin in the intes-
tine was only a third of that seen in liver.
To study topological differences between rat gut segments
in ability to metabolize curcumin, extent of metabolism in
cytosol and microsomes was compared between rat jejunum
and colon. The amounts of curcumin sulfate generated by
jejunal or colonic cytosol were 25 ⫾ 3 and 40 ⫾ 9 nmol/mg
protein (mean ⫾ SD, n ⫽ 3), respectively. Reductive metabo-
Fig. 3. On-line HPLC-mass spectrometry analysis in selected ion registration lism in cytosol from the jejunum furnished 8 ⫾ 3 nmol hexa-
mode of curcumin sulfate (molecular ion at m/z ⫽ 447) derived from extracts of hydrocurcumin/mg protein, whereas the amount of hexahydro-
incubations of curcumin with human intestinal cytosol (A) and recombinant curcumin detected in colonic cytosol was below the limit of
SULT1A1 (B). Arrow, retention time of authentic curcumin sulfate. For details of
incubation, extraction, and HPLC analysis, see “Materials and Methods.” The
quantification. Microsomes from the jejunum or colon gener-
chromatograms are representative of analyses conducted with cytosol from 2 ated 16 ⫾ 1 and 42 ⫾ 11 nmol curcumin glucuronide/mg
humans and two enzyme preparations each. protein, respectively (mean ⫾ SD, n ⫽ 3).
Metabolism of Curcumin by Intact Rat Gut. To explore the
metabolism of curcumin in an intact intestinal tissue, the agent
min generated as much as 16 times more curcumin glucuronide was incubated with inverted rat gut sacs. Curcumin sulfate was
in the intestine of humans than in the equivalent tissue in the rat, unequivocally identified by cochromatography and mass spec-
but human liver microsomes generated only a third of the trometry (result not shown). Curcumin glucuronide and prod-
amount of curcumin glucuronide found in rat liver microsomes. ucts of metabolic curcumin reduction were not detected.
Microsomal glucuronidation of curcumin in human intestine Nature and Location of Curcumin SULTs and Reductases.
exceeded that seen in liver by a factor of 2.5, whereas in the rat, The enzymes SULT1A1 and 1A3 are among five isoenzymes of
the amount of curcumin glucuronide formed in the intestine was the human phenol xenobiotic-metabolizing SULT subfamily,
only a 60th of that measured in the liver. There was consider- which are expressed in the gastrointestinal tract (30). To ex-
able variation in formation of curcumin glucuronide in human plore whether they may be involved with the generation of
intestine, perhaps related to interindividual differences in the curcumin sulfate in tissue fractions, curcumin was incubated
abundance of UDP-glucuronosyl-transferase enzymes influ- with recombinant SULTs. Analysis of extracts of the incubates
enced by the donor’s genotype, disease state, and lifestyle. by HPLC-mass spectrometry confirmed that SULT1A1 (Fig. 3)
Reduction of curcumin to hexahydrocurcumin was 18 times and SULT1A3 (result not shown) metabolize curcumin to its

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 Cancer Epidemiology, Biomarkers & Prevention 109

Fig. 4. Western blot analyses of human intestinal and hepatic tissue samples
with monoclonal antibodies against phenoltransferase isoenzymes SULT1A1 and
1A3. Assignation of lanes is as follows: A, recombinant SULT1A3; F, recom-
binant SULT1A1; B, C, and D, samples of intestine from 3 humans; E, sample of
liver from 1 human. Arrows, positions of the SULT1A3 (solid arrow) and
SULT1A1 (broken arrow) bands; in B–E, the top bands are indistinguishable
from that of SULT1A3, and the bottom bands comigrate with SULT1A1. For
details of Western analysis, see “Materials and Methods.” The blots are repre-
sentative of two separate experiments.

sulfate. At a curcumin substrate concentration of 100 ␮M, 6 Fig. 5. High-performance liquid chromatograms of extracts of incubations of
nmol curcumin sulfate/␮g protein were generated by SULT1A1 hexahydrocurcumin (A) or curcumin (B) with human intestinal microsomes.
during the 30-min incubation (mean of n ⫽ 2), and SULT1A3 Incubation time was 90 min, and analysis was conducted with detection at 280
catalyzed the production of four times this amount of curcumin nm. Peak allocation is hexahydrocurcumin (peak 1), hexahydrocurcuminol (6),
and curcumin (3). The identity of peak 6 as hexahydrocurcuminol was established
sulfate. Western blot analysis corroborated the presence of both by cochromatography with authentic material and mass spectrometry. Note that
SULT1A1 and 1A3 in human intestinal and hepatic cytosol commercially available curcumin contains 15% desmethoxycurcumin and 5%
(Fig. 4). bisdesmethoxycurcumin, which furnished two small peaks just beyond curcumin
To find out if alcohol dehydrogenases may be involved in (in B). AU, absorbance units. For details of extraction and HPLC analysis, see
“Materials and Methods.” The chromatograms are representative of two experi-
curcumin reduction, curcumin was incubated with horse alco- ments.
hol dehydrogenase. Hexahydrocurcumin was identified as a
metabolite by HPLC and mass spectrometry (result not shown).
Both hexahydrocurcumin and hexahydrocurcuminol have been
reported to be the predominant products of metabolic reduction capable of conjugating curcumin than the jejunum, at least in
of curcumin in incubations of intact human or rat hepatocytes the rat. The data shown in Table 1 allow a comparison to be
(18), whereas in the experiments with liver or gut cytosol made between humans and rats as to the capability of intestinal
described above, hexahydrocurcumin and tetrahydrocurcumin and hepatic tissues to metabolize curcumin, thus helping define
were the major curcumin reduction products, and only traces of the suitability of the rat as a model to study the metabolism of
hexahydrocurcuminol were found. To rationalize this differ- curcumin in humans. Whereas the pattern of metabolites of
ence in metabolism of curcumin between cellular cytosol on the curcumin in human intestinal and hepatic tissues was qualita-
one side and intact cells on the other, curcumin or hexahydro- tively similar to that in rat tissues, there were considerable
curcumin was incubated with human gut microsomes, fortified quantitative differences. In human intestinal fractions, conju-
with NADPH-generating cofactors. Fig. 5 shows that micro- gation of curcumin with activated sulfuric or glucuronic acids
somes reduced hexahydrocurcumin to hexahydrocurcuminol was much more abundant, whereas conjugation in human he-
but did not metabolize curcumin. This result means that patic fractions was much less extensive, than in rat tissues.
whereas curcumin is a poor substrate of human microsomal Furthermore, the ability of either intestinal or liver tissues from
reducing enzymes, hexahydrocurcumin is a good substrate of humans to reduce curcumin exceeded that in tissues from rat by
these enzymes. In contrast, human gut cytosol reduced curcu- factors of 18 and 5, respectively. These differences may reflect
min easily (see Fig. 2), and it also reduced hexahydrocurcumin discrepancies in tissue enzyme content. Taken together, these
to hexahydrocurcuminol (result not shown). results suggest that experiments in the rat may severely under-
estimate the extent of intestinal metabolism of curcumin, which
occurs in humans, a conclusion which hints at the possibility
Discussion that, in quantitative terms, the rat may not be a good model for
These are the first results to provide convincing evidence that the elucidation of the extrahepatic metabolic disposition of
curcumin is biotransformed in the intestinal tract of humans and curcumin in humans.
rodents. Metabolism of curcumin to curcumin glucuronide, The results also provide preliminary insights into the en-
curcumin sulfate, tetrahydrocurcumin, and hexahydrocurcumin zymology associated with intestinal metabolism of curcumin.
was demonstrated in intestinal fractions from humans and rats, Curcumin was shown to be a substrate of both phenol SULTs
and its conversion to curcumin sulfate was demonstrated in situ tested here, SULT1A1 and SULT1A3. Of the two, the latter
in intact rat intestine. These findings confirm and extend the was more efficient in sulfating curcumin. Curcumin sulfate is
original observation that in rat gut sacs, in situ [3H]labeled the first curcumin metabolite that has been identified in human
curcumin underwent metabolic removal from the incubation feces (19), and its generation may have been catalyzed by these
medium (23). They also corroborate the finding that intestinal enzymes in the gut. Alcohol dehydrogenase was pinpointed
mucosa, as well as liver and kidney tissue from the rat, can here as a potential source of metabolically generated hexahy-
glucuronidate and sulfate curcumin, as adjudged by analysis of drocurcumin. Nevertheless, it is probable that a variety of other
differential amounts of curcumin present before and after treat- ubiquitous and nonspecific oxido-reductases reduce curcumin
ment of tissue extracts with conjugate-hydrolyzing enzymes and thus contribute to the formation of curcumin reduction
(21). The results of the quantitative evaluation of curcumin products in vivo. The potential involvement of alcohol dehy-
metabolism in human and rat tissue fractions presented in Table drogenase enzymes with the cytosolic reduction of curcumin in
1 suggest that gut metabolism contributes substantially to the gut and liver potentially raises the clinically pertinent question
overall metabolite yield generated from curcumin in vivo. Fur- of whether the metabolic disposition of curcumin may be com-
thermore, they support the notion that the colon may be more promised by consumption of alcoholic beverages and, vice

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110 Intestinal Metabolism of Curcumin

(18). On balance, these findings justify the tentative conclusion

that the intestinal biotransformation of curcumin constitutes a
pharmacological deactivation process, in that metabolism gen-
erates species that are either devoid of biological activities
germane to cancer chemoprevention or less potent than their
metabolic precursor.
In conclusion, this study demonstrates that curcumin is
avidly metabolized by human intestinal tissue. The pharmaco-
logical implications of intestinal conjugation and bioreduction
of curcumin should be considered in the design of future cancer
chemoprevention trials of this interesting dietary constituent.

Fig. 6. Schematic representation of metabolism of curcumin in intestinal and Acknowledgments

hepatic cytosol (open arrows) and microsomes (closed arrows). We thank the United Kingdom MRC for financial support (MRC Toxicology Unit
core funding and studentship to C. R. I.), Dr. W. Wang (Phytopharm plc, Cam-
bridge, United Kingdom) for the synthesis and provision of tetrahydrocurcumin
and hexahydrocurcumin, the United Kingdom Human Tissue Bank for provision
versa, that dehydrogenation of dietary alcohol may be impaired of human tissue, and Drs. R. Sharma, R. Verschoyle, and P. Forshaw and U.
in individuals who consume large amounts of curcumin. Zafar, S. Oustric, L. Howells, and K. Hill for help with experiments and useful
discussions concerning the manuscript.
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Metabolism of the Cancer Chemopreventive Agent Curcumin
in Human and Rat Intestine
Christopher R. Ireson, Donald J. L. Jones, Samantha Orr, et al.

Cancer Epidemiol Biomarkers Prev 2002;11:105-111.

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